Phagocyte Contact Response in Aspergillosis Immunity

描述（由申请人提供）：烟曲霉是免疫功能低下人群中主要的空气传播真菌病原体。由于难以控制烟曲霉感染，大多数发展为侵袭性曲霉病的个体将死亡。随着免疫功能低下人群的增加，烟曲霉感染也会增加。虽然在了解曲霉病方面取得了重大进展，但对相应免疫细胞之间的关系尚未完全了解。这项工作的长期目标是确定寄主防御烟曲霉的分子基础。具体的重点是确定吞噬细胞与烟曲霉之间的细胞和分子接触以及杀微生物反应在相应免疫中的作用。为了实现这一目标，提出了三个目标：1)利用流式细胞术和免疫组织化学技术评估小鼠肺部对烟曲霉分生孢子虫的细胞特异性反应：将使用特异性单克隆抗体探测正常小鼠和免疫抑制小鼠接种烟曲霉分生孢子虫后到达的肺组织。将对可的松诱导的免疫抑制和敲除NADPH氧化酶使小鼠易感进行研究。2)为了研究NADPH氧化酶在摄入分生孢子的免疫细胞亚细胞区室中的作用：超氧化物生成试验将用于显示氧化酶与含有摄入分生孢子的吞噬溶酶体相关的组装。组装的复合物将通过在含有分生孢子的隔室中展示超氧化物的生成速率来证实，并通过免疫印迹证明氧化酶组分的完整集合。一组单克隆抗体可用于氧化酶检测。3)通过噬菌体展示肽库分析，鉴定出烟曲霉分生孢子和菌丝形态特异性结合的独特肽。利用肽库探测烟曲霉静息分生孢子、肿胀分生孢子和菌丝，进行独特肽的亲和选择。这些信息将用于提示寄主-病原体关系中的附着机制

DESCRIPTION (provided by applicant): Aspergillus fumigatus is the leading airborne fungal pathogen in immune compromised people. Due to difficulties in managing A. fumigatus infections, most individuals that develop invasive aspergillosis will die. With an increasing immune compromised population, infections by A. fumigatus will also increase. While significant advancements in understanding aspergillosis have been made, the relationship among corresponding immune cells is not fully understood. The long-term goal of this work is to identify the molecular basis of host defense against A. fumigatus. The specific focus is to determine the cellular and molecular contact between phagocytes and A. fumigatus and the role of microbicidal responses in the corresponding immunity. Three aims are proposed to address this goal: 1) To evaluate the cell specific response to A. fumigatus conidia in the lungs of mice by flow cytometry and immunohistochemistry: Lung tissue will be probed using specific monoclonal antibodies to identify cells that arrive following inoculation of A. fumigatus conidia in normal and immune suppressed mice. Both cortisone-induced immune suppression and mice made susceptible to by knockout of the NADPH oxidase will be examined. 2) To investigate the role of the NADPH oxidase in subcellular compartments of immune cells which have ingested conidia: Superoxide generation assays will be used to show assembly of the oxidase associated with phagolysosomes containing ingested conidia. The assembled complex will be confirmed by demonstrating superoxide generation rates in compartments containing conidia, as well as demonstrating the complete set of oxidase components by immunoblot. A panel of monoclonal antibodies is available for oxidase detection. 3) To identify unique peptides that bind specifically to the A. fumigatus conidia and hyphal forms by phage-display peptide library analysis. Affinity selection of unique peptides will be carried out by probing the A. fumigatus resting conidia, swollen conidia, and hyphae with a peptide library. This information will be used to suggest mechanisms of attachment in this host-pathogen relationship